The zebrafish, Danio rerio, has become widely accepted as a useful model for vertebrate development. There is strong interest in developing zebrafish as a model organism for use in high throughput drug discovery assays. Thus far, true high throughput screening in zebrafish has been limited to embryos and very early larvae. This is because many of the organs that are most interesting to researchers are best viewed from the side of the animal. Once the swim bladder inflates and the larvae generally float upright it is difficult to generate a clear image of the side of the animal. This is especially true in multiwell microplate format. Efforts to resolve this important problem have centered on use of confocal imaging techniques or powerful image deconvolution software to assemble 3-D reconstructions of the animal. These efforts have not achieved the desired spatial resolution and typically require 1 to 2 minutes to image a single well. Physical Sciences Inc. (PSI) has invented a novel technology that will be adaptable to standard image analysis systems that are already in use in drug discovery laboratories. PSI's technology will allow researchers to generate clear side-views of older zebrafish larvae in microwell arrays. The technology does not require confocal microscopy techniques or sophisticated image deconvolution algorithms and will therefore generate quality images in seconds per well versus minutes per well for those other techniques. In Phase I we used the Z-Lipotrack assay under development at Zygogen Corp to demonstrate the feasibility of our technology. The Lipotrack assay was chosen because it is typically performed on older larvae and requires collection of images of the gall bladder and intestine. In Phase II we will further develop PSI's innovative imaging technology and integrate that technology into a fully integrated, automated discovery system for zebrafish larvae. PSI's technology will enable high throughput discovery using the Z-Lipotrack assay of bioactive compounds that reduce cholesterol and lipid levels. Those drugs meet an important health care need. Relevance to public health: This research will develop technologies that will enable the use of zebrafish models of human disease in high-throughput screens for active pharmaceutical compounds. The specific assay under development is expected to accelerate discovery of cholesterol-lowering drugs. We anticipate that assays utilizing the imaging technology will also accelerate discovery of new Pharmaceuticals to treat a broad range of human illness such as cardiovascular disease and acute renal failure. [unreadable] [unreadable] [unreadable]